Evaluation of Aceclofenac Loaded Alginate Mucoadhesive Spheres Prepared by Ionic Gelation

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1 Varma and Rao: Evaluation of Aceclofenac Loaded Alginate Mucoadhesive Spheres Prepared by Ionic Gelation 1847 International Journal of Pharmaceutical Sciences and Nanotechnology Volume 5 Issue 4 January March 2013 Research Paper MS ID: IJPSN VARMA Evaluation of Aceclofenac Loaded Alginate Mucoadhesive Spheres Prepared by Ionic Gelation M.M. Varma* and Ch. H.L.N. Rao Shri Vishnu college of Pharmacy, Vishnupur, Bhimavaram , Andhra Pradesh, India. Received June 1, 2012; accepted July 14, 2012 ABSTRACT Controlled release aceclofenac spheres were prepared in a cross-linked alginate matrix using ionotropic gelation technique. A suspension of aceclofenac in sodium alginate solution was added drop wise into 10% w/v calcium chloride solution and the resulting spheres were evaluated for their drug content, flow properties, mucoadhesive property and the dissolution rate. The aceclofenac loaded alginate spheres were prepared using various mucoadhesive polymers: sodium alginate, HPMC, sodium CMC, guar gum, methyl cellulose and carbopol. The calcium chloride was used as the crosslinking agent. Fourier transform infrared spectroscopy (FTIR) was used to evaluate the drug-polymer interaction. The alginate spheres showed good rheological properties, drug content KEY WORDS: Aceclofenac, Mucoadhesive, Alginate spheres, Controlled release uniformity and high entrapment efficiency. The aceclofenac release from the spheres was slow and extended up to 10 hours. The drug loaded spheres exhibited good mucoadhesive property in the in vitro wash off test. The drug release from the optimized formulation (drug-sodium alginate-hpmc K15M; 1:0.9: 0.1 ratio) followed zero order kinetics and exhibited non- Fickian diffusion. The rate of release of aceclofenac decreased with increasing concentration of sodium alginate due to slow penetration of dissolution fluid in the spheres. The results suggest that alginate spheres can potentially deliver aceclofenac at zero-order controlled release following oral administration. The FTIR studies indicated the absence of the drug-polymer interaction in the solid state. Introduction Mucoadhesion is the relatively new and emerging concept in drug delivery. Mucoadhesion keeps the delivery system adhering to the mucus membrane. Mucoadhesion or the attachment of a natural or synthetic polymer to a biological substrate is a practical method of drug immobilization and an important new aspect of controlled drug delivery (Peppas and Bury, 1985; Castellanos et al., 1993; Ponchel and Irache, 1998). The primary objective of a mucoadhesive dosage form is to provide intimate contact of the dosage form with the absorbing surface and to increase the residence time of the dosage form at the absorbing surface and prolong the drug action. A number of mucoadhesive based dosage forms, including sustained release tablets, semisolid forms, powders, granules, micro- and nanoparticles in the GI tract have been widely studied (Bodde et al., 1996; Chowdary and Srinivas, 2000; Vasir et al., 2003; Patil and Murthy, 2006). Nonetheless, successful systems that will be retained in the GI tract of human for a desirable time have not yet been developed. Mucoadhesion also increases the intimacy and duration of contact between a drug-containing polymer and a mucous surface. The effect of prolonged residence time allows for an increased bioavailability of the drug with a smaller dosage and less frequent administration (Chowdary and Rao, 2003; Patel et al., 2005; Belgamwar et al., 2009).The mucoadhesive polymers containing carboxylic group exhibit the best mucoadhesive properties: sodium carboxy methyl cellulose (SCMC), hydroxyl propyl cellulose (HPC) and other cellulose derivatives (Das and Mourya, 2008; Patil et al., 2009; Thorat et al., 2009). Multi-particulate drug delivery systems have been widely used to provide controlled release of drugs because they exhibit many advantages over conventional single unit dosage forms (Patel et al., 2010). Some of these advantages include less inter- and intra-subject variability because their absorption is less dependent on gastric emptying and gastrointestinal transit time, greater bioavailability because granules or pellets disperse freely in the gastrointestinal tract, and decreased local irritation because of small amount of drug present in each unit. Therefore, multiparticulate drug delivery system in the form of spheres was selected as the formulation of choice. Several methods of preparing multi-particulate drug delivery systems are available, e.g., spheronization, spray granulation, and fluidized bed granulation. The main disadvantages associated with these techniques include high cost of manufacturing and requirement of specialized equipments (Patel et al., 2010). Therefore, ionic gelation technique was selected for preparation of controlled release spheres. This technique involves ionic cross- 1847

2 1848 Int J Pharm Sci Nanotech Vol 5; Issue 4 January March 2013 linking between polyelectrolyte biopolymers such as alginates, carboxymethylcellulose and chitosan. and counter ions to produce cross-linked matrix which would provide the desired controlled release rate (Patil et al., 2010). Sodium alginate was chosen as the biopolymer because alginates are non-toxic, biodegradable, naturally occurring anionic polysaccharides having high biological safety, and can absorb times their own weight of water (Almeida and Almeida, 2004). One of the most important and useful property of alginates is their ability to form gels in the presence of metal ions such as calcium. Sodium alginate is water-soluble and can be cross-linked with divalent or polyvalent cations to form insoluble meshwork. It can be easily gelled by the addition of calcium ions to form an insoluble calcium alginate network. This occurs due to cationic exchange between Na + and Ca 2+ ions and the technique is called ionotropic gelation (Kook and Jin, 1992). The alginate matrix obtained possess many advantages including nontoxicity, protection of mucous membrane from irritation upon oral administration, controlled release of drug due to its ability to undergo swelling when it comes in contact with water, and ability to incorporate acid sensitive drugs into the matrix (Almeida and Almeida, 2004). The non-steroidal anti-inflammatory drugs (NSAIDs) are being widely used in the treatment of osteoarthritis, rheumatoid arthritis and ankylosing spondylitis. Aceclofenac is an effective analgesic and antiinflammatory agent (Brogden and Wiseman, 1996). Aceclofenac is practically insoluble in water (0.3 mg/ml at 37⁰C). The mode of action of aceclofenac is largely based on the inhibition of prostaglandin synthesis. Aceclofenac is a potent inhibitor of the enzyme cyclooxygenase, which is involved in the production of prostaglandins. As the drug has low elimination half-life (4 hrs), mucoadhesive spheres were prepared to extend the release of the drug. An obstacle to the more successful use of aceclofenac therapy is the high incidence of GI symptoms seen in about 30% patients, especially during initial weeks of treatment. Patient compliance decreases with frequent dosing regimen and the side effects are associated with the same. The above drawbacks provide a rationale for developing aceclofenac as the mucoadhesive spheres, which will stick to the mucous membrane of GI tract and produce a constant input of drug to the absorption site. This improves the bioavailability of the drug, reduces frequency of dosing, minimizes side effects and enhances the patient compliance (Sarfaraz et al., 2010). Therefore, the objectives of this study were to investigate the feasibility of formulating controlled release aceclofenac loaded alginate spheres with a view to extend the drug release, enhance its oral bioavailability and efficacy. Materials and Methods Drugs and chemicals Aceclofenac was obtained as a gift sample from SK Parenterals, Tanuku. Carbopol 940P, HPMC K15M, HPMC K100M, HPMC 100 cps, ethyl cellulose, guar gum, methyl cellulose, sodium alginate, sodium carboxymethylcellulose (Na CMC) and calcium chloride were procured from S.D.Fine Chem. Ltd., Mumbai, India. All other chemicals used were of analytical grade. Formulation and Evaluation Methodology Estimation of aceclofenac A spectrophotometric method (Chowdary and Rao, 2003) based on the measurement of absorbance at 273 nm in ph 7.4 phosphate buffer was used in the present study for the estimation of aceclofenac. The 100 mg of aceclofenac pure drug was dissolved in 20 to 30 ml of alcohol, then the volume was adjusted to 100 ml with ph 7.4 phosphate buffer (stock solution 1000 μg/ml), from this 10 ml of solution was taken and the volume was adjusted up to100 ml with ph 7.4 phosphate buffer (100 μg/ml). The above solution was subsequently diluted with ph 7.4 phosphate buffer to obtain the series of dilutions containing 2, 4, 8, 12, 16, 20, 24 and 30 μg/ml of aceclofenac solution. The absorbance of the above dilutions was measured at 273 nm by using the UVspectrophotometer (Analytical) taking ph 7.4 phosphate buffer as the blank. Then a graph was plotted by taking concentration on x-axis and absorbance on y-axis which gives a straight line. Preparation of mucoadhesive spheres All the formulations were prepared by orifice ionic gelation method (Kim and Lee, 1992; Hari et al., 1996). The composition of different formulations is given in Table 1. The alginate spheres were prepared as per the procedure given below: Aceclofenac and all other polymers were individually passed through sieve # 60. Sodium alginate (1.0 g) and the mucoadhesive polymer (1.0 g) were dissolved in purified water (32 ml) to form a homogenous polymer solution. The active substance, Aceclofenac (1.0 g), was added to the polymer solution and mixed thoroughly with a stirrer to form a viscous dispersion. The resulting dispersion was then added manually drop wise into calcium chloride (10 % w/v) solution (40 ml) through a syringe with a needle of size no. 18. The added droplets were retained in the calcium chloride solution for 15 minutes to complete the curing reaction and to produce spherical rigid drug loaded spheres. The spheres were collected by decantation, and the product thus separated was washed repeatedly with water and dried at 45 ⁰C for 12 hours. Evaluation of mucoadhesive spheres The formulated alginate spheres were evaluated for the following physicochemical characteristics: Angle of repose Angle of repose is defined as the maximum angle possible between the surface of the pile of the powder and the horizontal plane (Patel et al., 2005). Angle of repose of different formulations was measured according to fixed height funnel standing method, tan θ = h / r,θ =

3 Varma and Rao: Evaluation of Aceclofenac Loaded Alginate Mucoadhesive Spheres Prepared by Ionic Gelation 1849 tan 1 (h / r). Where, h = height of pile, r = radius of the base of the pile, θ = angle of repose. Bulk density and tapped density Bulk density and tapped density were measured by using 10 ml of graduated cylinder. The sample poured in the cylinder was tapped mechanically for 100 times, and then the tapped volume was noted down. Bulk density and tapped density were calculated (Patel et al., 2005). Each experiment for micromeritic properties was performed in triplicate. Bulk Density = (Weight of the microspheres / Bulk volume). Tapped Density = (Weight of the microspheres / Tapped volume). Carr s index Carr s index value of the alginate spheres was computed according to the following equation: Carr index (%) = [(Tapped density Bulk Density) / Tapped Density] 100 Hausner s ratio Hausner s ratio of the drug loaded spheres was determined by estimating the ratio of the tapped density to the bulk density (Patel et al., 2005), using the equation: Hausner s Ratio = Tapped density / Bulk Density. Drug content Preparations equivalent to 50 mg of drug was weighed accurately and transferred to a 100 ml volumetric flask and dissolved in 20 ml of alcohol. The volume was adjusted with ph 7.4 phosphate buffer. The sample was filtered and after suitable dilution, the absorbance of the above solution was measured at 273 nm. The drug content was calculated using the calibration curve (Chowdary and Rao, 2003). Size analysis For size distribution analysis, different sizes in a batch were separated by sieving using a set of standard sieves. The amounts retained on different sieves were weighed (Goudanavar et al., 2010). Entrapment efficiency Entrapment efficiency was calculated using the following formula (Sarfaraz et al., 2010): entrapment efficiency = (estimated percentage drug content/ theoretical percentage drug content) 100. Mucoadhesion testing by ex vivo wash-off test The mucoadhesive property of the aceclofenac loaded spheres was evaluated by an in vitro mucoadhesion testing method known as the in vitro wash-off method (Rao et al., 1998; Chowdary and Rao, 2003). Freshly excised pieces of intestinal mucosa (4 4 cm) from the sheep were mounted on to glass slides (3 1 inch) with cyanoacrylate glue. Two glass slides were connected with a suitable support. About 100 spheres were spread on to each wet rinsed tissue specimen, and immediately there after the support was hung on to the arm of a USP tablet disintegrating test machine. When the disintegrating test machine was operated, the tissue specimen was given a slow, regular up and down movement in the test fluid at 37 C contained in a 1 L vessel of the machine. At the end of 30 minutes, at the end of 1 hour, and at hourly intervals up to 10 hours, the machine was stopped and the number of spheres still adhering to the tissue was counted. The test was performed at intestinal ph (7.4 phosphate buffer). Dissolution study The 900 ml of ph 7.4 phosphate buffer was placed in the vessel and the USP apparatus II (paddle method) was assembled (Chowdary and Rao, 2003). The medium was allowed to equilibrate to temperature of 37 C C. The drug loaded spheres were placed in the vessel and the vessel was covered, the apparatus was operated for 10 hrs at 50 rpm. At definite time intervals, the 5 ml of the dissolution fluid was withdrawn; filtered and again 5ml blank sample was replaced. The samples were analyzed spectrophotometrically at 273 nm (λ max) using a UV-spectrophotometer (Analytical). Release kinetics The analysis of drug release mechanism from a pharmaceutical dosage form is an important but complicated process and is practically evident in the case of mucoadhesive controlled release systems. The order of drug release from the mucoadhesive spheres was described by using zero order kinetics or first orders kinetics (Wamorkar et al., 2010; Varma and Vijaya, 2012). The mechanism of drug release from the mucoadhesive controlled release spheres was studied by using the Higuchi equation (Higuchi, 1963) and the Korsemeyer Peppas equation (Peppas, 1985). Zero order release kinetics It defines a linear relationship between the fraction of drug released versus time. Q = kot. Where, Q is the fraction of drug released at time t and ko is the zero order release rate constant. A plot of the fraction of drug released against time will be linear if the release obeys zero order release kinetics. First order release kinetics Wagner assuming that the exposed surface area of a tablet decreased exponentially with time during dissolution process suggested that drug release from most of the slow release tablets could be described adequately by apparent first-order kinetics. The equation that describes first order kinetics is: In (1-Q) = - K1t, where, Q is the fraction of drug released at time t and K1 is the first order release rate constant. Thus, a plot of the logarithm of the fraction of drug undissolved against time will be linear if the release obeys the first order release kinetics. Higuchi equation It defines a linear dependence of the active fraction released per unit of surface (Q) and the square root of time: Q=K2t ½, where K2 is the release rate constant. A

4 1850 Int J Pharm Sci Nanotech Vol 5; Issue 4 January March 2013 plot of the fraction of drug released against square root of time will be linear if the release obeys Higuchi equation. This equation describes drug release as a diffusion process based on the Fick s law, square root time dependant. Power law In order to define a model, which would represent a better fit for the formulation, dissolution data was further analyzed by Peppa s and Korsemeyer equation (Power Law), Mt/M α = K.t n. Where, Mt is the amount of drug released at time t and M α is the amount released at time α, thus the Mt/M α is the fraction of drug released at time t, K is the kinetic constant and n is the diffusion exponent..a plot between log of Mt/M α against log of time will be linear if the release obeys Peppa s and Korsemeyer equation and the slope of this plot represents n value. FTIR studies FTIR spectrum of the drug, polymers, and the drugpolymer physical mixture were studied to confirm the compatibility of the drug with the polymer (Varma and Vijaya, 2012). FTIR spectra were recorded by using the FTIR spectrophotometer (Bruker) using KBr pellets and the scanning range used was 4400 to 400 cm -1 at a scan period of 1 min. Similarity factor (f2 analysis) f2 = 50. log { [1 + ( 1/n) t=1 n (Rt - Tt ) 2 ] } Where 'Rt' and Tt' are the cumulative percentage drug dissolved at each of the selected n time point of the reference and test product respectively. The factor f2 is inversely proportional to the averaged squared difference between the two profiles, with emphasis on the larger difference among all the time points (Varma and Vijaya, 2012). Results and Discussion Preparation of mucoadhesive spheres In the present study mucoadhesive spheres of aceclofenac were prepared to reduce the side effects like gastric irritation, to increase the drug bio-availability, extend the drug release, to reduce the frequency of dosing, and to enhance the patient compliance. The alginate spheres were prepared by orifice-ionotropic gelation method using the mucoadhesive polymers such as HPMC (K 15 M, K 100 M, 100 cps), Carbopol 940, Na CMC, Guar gum, Sodium Alginate and Methyl Cellulose. The 10% Calcium Chloride solution was used as a crosslinking agent (Chowdary and Rao, 2003). The sodium salt of aceclofenac was not used in this investigation because sodium ions on the drug molecule could preferentially exchange calcium ions in calcium chloride solution leading to poor or no cross-linking of sodium alginate with calcium chloride. Crosslinking between alginate and calcium ions may lead to ionotropic gelation (Arica et al., 2005). Since the size of a droplet formed at the end of a needle depends on the size of needle opening, a needle with 18 gauge size was found to be suitable for preparing the spheres. Needles having larger openings produced larger sized spheres which were not acceptable for this study and needles with smaller openings made the flow of naproxen-alginate suspension more difficult. Immediately after separation from calcium chloride solution, the alginate spheres were found to be very sticky to touch (Khazaeli et al., 2008). The texture was smooth, but the spheres appeared soft, indicating that the interaction with calcium chloride may not have hardened the alginate spheres sufficiently (Bajpai and Sharma, 2004). Upon drying, they shrunk as well as became relatively harder. Totally, 16 different formulations of aceclofenac were prepared by using the above polymers (Table 1). Finally, the spheres were evaluated for various characteristics like drug content, entrapment efficiency, percent mucoadhesive strength and the in vitro release was evaluated for 10 hrs. TABLE 1 Composition of aceclofenac loaded spheres. S No Batch code Coat Composition Ratio 1 F1 Drug: Sod. Alginate:Ethyl cellulose 1:1:1 2 F2 Drug: Sod. Alginate : Sodium CMC 1:1:1 3 F3 Drug: Sod. Alginate 1:1 4 F4 Drug: Sod. Alginate: Guar gum 1:1:1 5 F5 Drug: Sod. Alginate : Methyl cellulose 1:1:1 6 F6 Drug: Sod. Alginate:Carbopol 940 1:1:1 7 F7 Drug: Sod. Alginate : HPMC 100Cps 1:1:1 8 F8 Drug: Sod. Alginate : HPMC K15M 1:1:1 9 F9 Drug: Sod. Alginate : HPMC K100M 1:1:1 10 F10 Drug: Sod. Alginate : Ethyl cellulose 1:0.9: F11 Drug: Sod. Alginate:Sodium CMC 1:0.9: F12 Drug : Sod.Alginate : Guar gum 1:0.9: F13 Drug: Sod. Alginate : Methyl cellulose 1:0.9: F14 Drug: Sod. Alginate : Carbopol 940 1:0.9: F15 Drug: Sod. Alginate :HPMC K15M 1:0.9: F16 Drug: Sod. Alginate : HPMC K100M 1:0.9:0.1 Evaluation of mucoadhesive spheres The drug loaded spheres were found to be discrete, spherical, free-flowing, and of the monolithic matrix type. All the formulations demonstrated good flow property as the values of angle of repose, Hausner s ratio and the compressibility index of the prepared spheres were very low (Table 2). The values of angle of repose of the different formulations ranged from 11 to 18 o.the values of the bulk density of the different batches ranged from g/ml g/ml. The values of the tapped density of the different formulations ranged from 0.295g/ml g/ml. The values of the Hausner s ratio of the different formulations ranged from 1 to The values of the Carr s index of the different batches ranged from 0% to 14.3%.The spheres were uniform in size, the diameter of the spheres ranged from 920µ-1220µ.The % drug content,% entrapment efficiency and the % mucoadhesive property of the different batches of the spheres are indicated in Table 3. The entrapment efficiency was in the range of 68% to 86%(Table 3).The

5 Varma and Rao: Evaluation of Aceclofenac Loaded Alginate Mucoadhesive Spheres Prepared by Ionic Gelation 1851 spheres with a coat consisting of sodium alginate and a mucoadhesive polymer exhibited good mucoadhesive property in the ex vivo wash-off test (Belgamwar et al., 2009; Patil et al., 2009; Thorat et al., 2009). The % mucoadhesive property of the different batches of spheres ranged from 50%-75%. Based on the % mucoadhesive property, the various formulations can be arranged,as:f6=f15(75%)>f2=f8=f16(70%).>f1=f4=f9 =F11=F12=F14(65%)>F3=F10(60%)>F5(55%)>F7=F13(5 0%).The formulations: F6 and F15 showed the highest mucoadhesive property(75%). FTIR studies The FTIR spectrum of the drug, HPMC K15M, sodium alginate and the drug-polymer physical mixture are depicted in Fig.1-4. The FTIR spectrum of the pure drug (Fig.1) shows the characteristic peaks of secondary amine at cm -1, ester stretching at cm -1, C-H aromatic stretching at cm -1, C=C aromatic stretching at cm -1 and the carboxylic vibration at cm -1. The FTIR spectrum of the drug and the polymer physical mixture (drug:sodium alginate:hpmc K15M,1: 0.9: 0.1) exhibited all the peaks of the drug. This confirms the undisturbed structure of the drug in the formulation. This proves the fact that there is no potential incompatibility of the drug with the polymers used in the formulations. The FTIR spectra clearly indicated the lack of drug-polmer interaction (Varma and Vijaya, 2012), as all the important peaks of the drug were observed in the drug-polymer physical mixture. TABLE 2 Flow properties of different formulations (n=3 ± S.D.) Formulation Angle of Repose(θ) Bulk density (g/ml) Tapped Density (g/ml) Hausner ratio Compressibility index (%) F1 11± ± ± ± ± F2 13± ± ± ± ± F3 14± ± ± ± ± F4 16± ± ± ± ±0.040 F5 12± ± ± ± ±0.062 F6 12± ± ± ± ±0.074 F7 11± ± ± ± ± F8 13± ± ± ± ± F9 14± ± ± ± F10 12± ± ± ± ± F11 16± ± ± ± ± F12 18± ± ± ± ± F13 12± ± ± ± ± F14 17± ± ± ± ± F15 13± ± ± ± ± F16 13± ± ± ± ± TABLE 3 Quality control parameters of mucoadhesive spheres of aceclofenac. Formulation Percent Drug content Encapsulation efficiency (%) Percent Mucoadhesive property F ± ± F2 36.0± ± F ± ± F4 37.2± ± F ± ± F6 34.0± ± F7 38.4± ± F ± ± F ± ± F ± ± F ± ± F ± ± F ± ± F ± ± F ± ± F ± ±

6 1852 Int J Pharm Sci Nanotech Vol 5; Issue 4 January March 2013 Fig. 1 FTI R spectrum of pure drug (aceclofenac). Fig. 2 FTIR spectrum of HPMC K 15 M. Fig. 3 FTIR spectrum of sodium alginate.

7 Varma and Rao: Evaluation of Aceclofenac Loaded Alginate Mucoadhesive Spheres Prepared by Ionic Gelation 1853 Fig. 4 FTIR spectrum of physical mixture (Aceclofenac: Sodium alginate: HPMC K 15M, 1:0.9:0.1). Dissolution study Aceclofenac release from the drug loaded spheres was studied in the phosphate buffer (ph 7.4) for 10 hours. The dissolution profiles of the different batches of the spheres are depicted in Fig.5-8. The dissolution parameters of the different formulations are indicated in Table 5. Aceclofenac release from the spheres was slow and depended on the composition of the coat (Chowdary and Rao, 2003; Sarfaraaz et al., 2010). Based on the T50 (hrs) values (time taken for 50% of drug release), the drug release from the different formulations can be arranged as: F1=F2=F15>F6>F3>F11>F14=F16>F10>F5=F12>F4 >F8>F13>F7>F9. Similarly, based on the T75 (hrs) values (time taken for 75% of drug release), the drug release from the different batches can be arranged as: F15>F16>F6>F14>F12>F1>F13>F8>F2=F11>F10>F 3=F4>F5. The batches, F7 and F9 did not attain T75 values even after 10 hours of the dissolution study. The formulations: F15 and F16 achieved 90% of drug release in 7.5 hours. All other formulations did not attain T90 (hrs) values (time taken for 90 % of drug release) even after 10 hours of the dissolution study. The differences in the drug release characteristics of the various spheres are due to the differences in the porosity of the coat formed and its solubility in the dissolution fluid (Hari et al., 1996; Arica et al, 2005). Among all the batches, the F15 (Drug: Sod. Alginate: HPMC K15M = 1:0.9:0.1) batch is considered to be the optimized formulation (T50% = 1.5 h,t75% = 4.5 h, T90% = 7.5 h ), because among all the batches it shows better extent of drug release, good entrapment efficiency (86.12%), and the in vitro wash-off test showed good mucoadhesive property(75%). Aceclofenac release from the optimized formulation (F15) was slow and extended over a period of 10 hrs and these drug loaded spheres were found suitable for the oral controlled release (Chowdary and Rao, 2003; Sarfaraz et al., 2010). The similarity factor was calculated. The similarity factor calculated for the batch F15 is 65%, so its release profile is similar to the dissolution profile of the aceclofenac marketed extended release tablet. The initial release of aceclofenac appears to depend on the concentration of alginate in the spheres. Smaller concentration of alginate may have produced more porous spheres which released aceclofenac more quickly (Ponchel and Irache, 1998). Moreover, spheres containing smaller alginate concentration may have produced a relatively weaker network which broke down faster than the relatively stronger network formed in spheres containing a larger concentration of alginate (Thorat et al., 2009). In most release studies dealing with multiparticulate systems, an initial burst effect is reported due to migration of drug to the surface of the particles. In this investigation, a burst effect was exhibited by the spheres containing low concentration of the polymer. The initial slow release was followed by a linear rate of release until almost 90% of the drug release (Patil et al., 2009). The decrease in release rate of drug from the alginate spheres (compared to almost instantaneous dissolution of pure drug in ph 7.4 phosphate buffer) is because of increase in the degree of cross-linking with the increase in concentration of sodium alginate. As the degree of cross-linking increases, the porosity decreases and the reduced porosity will further retard the release of drug from the alginate spheres (Khazaeli et al., 2008). Also, drug release from a hydrophilic matrix is controlled by the formation of a hydrated viscous layer around the matrix which acts as a barrier to drug release by opposing penetration of water in to the matrix and also

8 1854 Int J Pharm Sci Nanotech Vol 5; Issue 4 January March 2013 movement of dissolved solutes out of the matrix (Patil et al., 2010). In case of spheres containing the higher concentration of the mucoadhesive polymer, the hydrophilic property of the polymer may bind better with water to form the viscous gel structure, which may block the pores on the sphere surfaces and prolong the drug release. As the polymer concentration of the prepared spheres was increased, the release rate was decreased. An inverse relationship was observed between the polymer concentration and the drug release from the prepared spheres (Patel et al., 2010). The release of drug was considered to occur mostly by diffusion but could be accelerated by the weight loss of the mucoadhesive polymers (Arica et al., 2005). The structure of the spheres changed significantly over time,indicating that there was substantial hydration and swelling of the mucoadhesive polymeric matrix. The alginatemucoadhesive polymeric gel might have acted as a barrier to the penetration of the dissolution medium, there by suppressing the diffusion of the drug from the swollen alginate-mucoadhesive polymeric matrix (Bolgamwaret al., 2009). The release of the drug was modulated by the diffusion of the drug through the swollen polymeric matrix (Gallardo et al., 1998). Fig. 5 Dissolution profile of mucoadhesive spheres of aceclofenac (F1, F2, F3, F4 formulations compared with brand). Fig. 6 Dissolution profile of mucoadhesive spheres of aceclofenac (F5, F6, F7, F8 formulations compared with brand).

9 Varma and Rao: Evaluation of Aceclofenac Loaded Alginate Mucoadhesive Spheres Prepared by Ionic Gelation 1855 Fig. 7 Dissolution profile of mucoadhesive spheres of aceclofenac (F9, F10, F11, F12 formulations compared with brand). Fig. 8 Dissolution profile of mucoadhesive spheres of aceclofenac (F13, F14, F15, F16 formulations compared with brand). Release Kinetics The regression coefficient (R 2 ) values of the different formulations are indicated in Table 4.Based on the values of R 2, the drug release from the formulations (F1, F2, F3, F4, F6, F7, F10, F11, F12, F14) exhibited first order kinetics, where as the drug release from the batches: F5, F8, F9, F13, F15, F16 demonstrated zero order kinetics. The values of the release rate constants, K (for the zero order, first order) and the diffusion exponent(n) are represented in Table 5. The values of K1(h -1 ), the first order release rate constant of the different batches ranged from 0.020(batch F9) to (batch F6). The values of K0(mg/h), the zero order release rate constant of the different formulations ranged from 2.70(F7) to 13.75(F3). All the formulations obeyed Higuchi equation (R 2 > 0.9), indicating that the drug release mainly depends on diffusion and erosion. The values of n (diffusion exponent) for all the formulations ranged from (batch F2) to 1.275(batch F8). Based on the n values, the drug release from the formulations: F1, F2, F6, F7, F11 followed Fickian diffusion(n<0.45), whereas the drug release from the batches: F3-F5, F10, F14, F15 exhibited non-fickian diffusion(n > 0.45),the drug release from the formulation F8, F9, F12, F13, F16 demonstrated super case-ii transport (n > 0.89) mechanism controlled by swelling and relaxation of the polymeric matrix. The drug release for the optimized formulation, F15, followed zero-order kinetics (Wamorkar et al., 2010; Varma and Vijaya, 2012) and the R 2 value was (Table 4). The Higuchi plot of F15 formulation showed an R 2 value of (Table 4), suggesting that the diffusion and erosion plays an important role in extending the drug release (Chowdary and Rao, 2003; Varma and Vijaya, 2012). The data was fitted to the Korsemeyer- Peppas equation and the value of diffusion exponent n for the batch F15 was 0.47 (Table 5) which indicated that the drug release (Chowdary and Rao, 2003) shows non- Fickian diffusion.

10 1856 Int J Pharm Sci Nanotech Vol 5; Issue 4 January March 2013 TABLE 4 Coefficient of regression (R 2 ) values of aceclofenac loaded spheres. Formulation Zero order First order Higuchi s Peppa s F F F F F F F F F F F F F F F F Brand Conclusions From the studies conducted in this investigation, it can be concluded that the release of aceclofenac can be retarded by encapsulating it in calcium alginate spheres. The optimized formulation, drug-sodium alginate-hpmc K15M (1: 0.9: 0.1 ratio) mucoadhesive spheres demonstrated controlled release of the drug for 10 hours and exhibited good mucoadhesive property. The FTIR studies revealed the absence of drug-polymer interaction in the solid state. The formulated mucoadhesive multiparticulate drug delivery system of aceclofenac can control the drug release; it has good mucoadhesive property and can improve the bioavailability of the drug. Acknowledgements The authors are grateful to the management of Shri Vishnu College of Pharmacy for providing the facilities to carry out this work. References Almeida FP and Almeida AJ (2004). Cross-linked alginategelatinbeads: a new matrix for controlled release of pindolol. J Control Release 97: Arica B, Calis S, Atilla P, Durlu NT, Cakar N, Kas HS and Hincal AA (2005). In vitro and in vivo studies of ibuprofen loaded biodegradable alginate beads. Journal of microencapsulation 22: Bajpai SK and Sharma S (2004). Investigation of swelling/ degradation behavior of alginate beads crosslinked with Ca 2+ and Ba 2+ ions. React Funct Polym 59: Belgamwar V, Shah V and Surana SJ(2009). Formulation and evaluation of oral mucoadhesive multiparticulate system containing metoprolol tartarate: An In Vitro Ex Vivo characterization. Current Drug Delivery 6: Bodde HE, De Vries ME and Junginger HE (1990). Mucoadhesive polymers for the buccal delivery of peptides, structure adhesiveness relationships. J Control Rel 13: Brogden RN and Wiseman LR (1996). Aceclofenac: A review of its pharmacodynamic properties and therapeutic potential in the treatment of rheumatic disorders and in pain management. Drugs 52: TABLE 5 Dissolution parameters of mucoadhesive spheres of aceclofenac. Formulation n K0 (mg/hr) Dissolution Parameters K1 (h -1 ) T50 (hrs) T75 (hrs) T90 (hrs) F >10 F >10 F >10 F >10 F >10 F >10 F >10 >10 F >10 F >10 >10 F >10 F >10 F >10 F >10 F >10 F F Brand Castellanos NRJ, Zia H and Rhodes CT (1993). Mucoadhesive drug delivery systems. Drug Dev Ind Pharm 19: Chowdary KPR and Srinivas L (2000). Mucoadhesive drug delivery systems. Indian Drugs 37: Chowdary KPR and Rao YS (2003). Design and In Vitro and In Vivo evaluation of mucoadhesive microcapsules of glipizide for oral controlled release. AAPS Pharm Sci Tech 4: Das MK and Mourya DP (2008). Evaluation of diltiazem hydrochloride-loaded mucoadhesive microspheres prepared by emulsification-internal gelation technique. Acta Poloniae Pharmaceutica Drug Research 65: Gallardo A, Eguiburu JL, Berridi MJF and Roman JS (1998). Preparation and in vitro release studies of ibuprofen loaded films and microspheres made from graft copolymers of poly(llactic acid) on acrylic back bones. J Control Rel 55: Goudanavar PSM, Patil S and Manvi FV (2010). Design and characterisation of sustained release microcapsules of salbutamol sulphate. International Journal of Pharm Tech Research 2: Hari PC, Chandy T and Sharma CP (1996). Chitosan/calcium alginate microcapsules for intestinal delivery of nitrofurantoin. J Microencapsulation 13: Higuchi T (1963). Mechanism of sustained-action medication. theoretical analysis of rate of release of solid drugs dispersed in solid matrices. J Pharm Sci 51: Khazaeli P, Pardakhtya A and Hassanzadeh F (2008). Formulation of ibuprofen beads by ionotropic gelation. Iranian J Pharm Res 7: Kim CK and Lee EJ (1992). The controlled release of blue dextran from alginate beads. Int J Pharm 79: Patel H, Patel JK and Patel RR (2010). Pellets: A general overview. Int J Pharm World Res 1: Patel JK,Patel RP, Amin AF and Patel MM(2005). Formulation and evaluation of mucoadhesive glipizide microspheres. AAPS Pharm Sci Tech 6: E49-E55. Patil JS, Kamalapur MV and Marapur SC (2010). Ionotropic gelation and polyelectrolyte complexation; the novel techniques to design hydrogel particulate sustained, modulated drug delivery system. Digest J Nanomat Biostru 5: Patil PB, Gawali VU, Patil HN, Hardikar SR and Bhosale AV (2009). Preparation and evaluation of mucoadhesive microspheres of atenolol and propranolol. Int J PharmTech Res 1: Patil SB and Murthy RSR (2006). Mucoadhesive polymers: means of improving drug delivery. Pharma Times 38:

11 Varma and Rao: Evaluation of Aceclofenac Loaded Alginate Mucoadhesive Spheres Prepared by Ionic Gelation 1857 Peppas NA (1985). Analysis of fickian and non-fickian drug release from polymers. Pharma Acta Helv 60: Peppas NA and Bury PA (1985). Surface interfacial and molecular aspects of polymer bioadhesion on soft tissues. J Control Rel 12: Ponchel G and Irache J (1998). Specific and non-specific bioadhesive particulate systems for oral delivery to the gastrointestinal tract. Adv Drug Deliv Rev 34: Rao YM, Vani G, Chary R and Bala R (1998). Design and evaulation of mucoadhesive drug delivery systems. Indian Drugs 35: Sarfaraz MD, Hiremath D and Chowdary KPR (2010). Formulation and characterization of rifampicin microcapsules. Indian J Pharm Sci 72: Thorat YS, Modi VS and Dhavale SC (2009). Use of carbomers to design mucoadhesive microspheres for an anti- H. pylori drug, clarithromycin. Int J PharmTech Research 1: Varma MM and Vijaya S (2012). Development and evaluation of gastroretentive floating drug delivery system of atenolol. Int J Pharm Chem Sci 1: Vasir JK, Tambwekar K and Garg S (2003). Bioadhesive microspheres as a controlled drug delivery system. Int. J Pharm 255: Wamorkar VV, Varma MM, Kumar BV and Reddy VM (2010). Effect of hydrophilic and hydrophobic polymers and in vitro evaluation of hydrodynamically balanced system of metoclopramide HCl. Int J Pharm Sci Nanotech 3: Address correspondence to: M. Mohan Varma, Shri Vishnu college of Pharmacy, Vishnupur, Bhimavaram , Andhra Pradesh, India. mohan_pharm@rediffmail.com